A novel predictive algorithm to personalize autologous T-cell harvest for chimeric antigen receptor T-cell manufacture.

BACKGROUND AIMS: The most widely accepted starting materials for chimeric antigen receptor T-cell manufacture are autologous CD3+ T cells obtained via the process of leukapheresis, also known as T-cell harvest. As this treatment modality gains momentum and apheresis units struggle to meet demand for harvest slots, strategies to streamline this critical step are warranted. METHODS: This retrospective review of 262 T-cell harvests, with a control cohort of healthy donors, analyzed the parameters impacting CD3+ T-cell yield in adults with B-cell malignancies. The overall aim was to design a novel predictive algorithm to guide the required processed blood volume (PBV) (L) on the apheresis machine to achieve a specific CD3+ target yield. RESULTS: Factors associated with CD3+ T-cell yield on multivariate analysis included peripheral blood CD3+ count (natural log, ×109/L), hematocrit (HCT) and PBV with coefficients of 0.86 (95% confidence interval [CI], 0.80-0.92, P < 0.001), 1.30 (95% CI, 0.51-2.08, P = 0.001) and 0.09 (95% CI, 0.07-0.11, P < 0.001), respectively. The authors' model, incorporating CD3+ cell count, HCT and PBV (L), with an adjusted R2 of 0.87 and root-mean-square error of 0.26 in the training dataset, was highly predictive of CD3+ cell yield in the testing dataset. An online application to estimate PBV using this algorithm can be accessed at https://cd3yield.shinyapps.io/cd3yield/. CONCLUSIONS: The authors propose a transferrable model that incorporates clinical and laboratory variables accessible pre-harvest for use across the field of T-cell therapy. Pending further validation, such a model may be used to generate an individual leukapheresis plan and streamline the process of cell harvest, a well-recognized bottleneck in the industry.

Re-evaluation of progenitor thresholds and expectations for haematopoietic recovery based on an analysis of 810 autologous transplants: Implications for quality assurance.

Haematological engraftment was assessed in 804 autologous transplants. Neutrophil recovery occurred in over 99% within 14 d but platelet recovery was delayed beyond this time in 14·8%. Time to recovery was dependent on the progenitor cell dose infused. The minimum CD34+ cell threshold adopted in this study (2 × 106 /kg) was safe although recovery was faster with a dose >5 × 106 /kg. CD34+ cell doses of between 1 and 2 × 106 /kg were also acceptable if either the granulocyte-macrophage colony-forming cell dose exceeded 2 × 105 /kg or this dose was due to splitting a higher yield harvest. Prompt neutrophil recovery affords important quality assurance for laboratory processing.

Post-thaw viability of cryopreserved peripheral blood stem cells (PBSC) does not guarantee functional activity: important implications for quality assurance of stem cell transplant programmes.

Standard quality assurance (QA) of cryopreserved peripheral blood stem cells (PBSC) uses post-thaw viable CD34(+) cell counts. In 2013, concerns arose at Great Ormond Street Hospital (GOSH) about 8 patients with delayed engraftment following myeloablative chemotherapy with cryopreserved cell rescue, despite adequate post-thaw viable cell counts in all cases. Root cause analysis was undertaken; investigations suggested the freeze process itself was a contributing factor to suboptimal engraftment. Experiments were undertaken in which a single PBSC product was divided into three and cryopreserved in parallel using a control-rate freezer (CRF) or passive freezing method (-80°C freezer) at GOSH, or the same passive freezing at another laboratory. Viable CD34(+) counts were equivalent and adequate in each. Granulocyte-monocyte colony-forming unit assays demonstrated colonies from the products cryopreserved using passive freezing (both laboratories), but no colonies from products cryopreserved using the CRF. The CRF was shown to be operating within manufacturer's specifications with freeze-profile within acceptable limits. This experience has important implications for quality assurance for all transplant programmes, particularly those using cryopreserved products. The failure of post-thaw viable CD34(+) counts, the most widely used routine QA test available, to ensure PBSC function is of great concern and should prompt reassessment of protocols and QA procedures.

Peripheral blood stem cell yield in 400 normal donors mobilised with granulocyte colony-stimulating factor (G-CSF): impact of age, sex, donor weight and type of G-CSF used.

Mobilised peripheral blood is now the main source of stem cells collected from normal donors. We report our experience of mobilising and collecting 400 normal healthy donors using standardised procedures and techniques. Target recipient doses were reached with one aphaeresis in 63% of donors and with two aphereses in 81% of donors. Approximately 2% of donors yielded such low progenitor values that they were termed 'poor mobilisers'. There were minor effects of donor age, weight and sex and where possible, larger male donors under the age of 55 years should be selected. Two forms of granulocyte colony-stimulating factor (G-CSF) were used at the same dose and no significant difference was seen in the yield of CD34+ cells collected/l of blood processed. However, a greater number of granulocyte-macrophage colony-forming cells were harvested using lenograstim (glycosylated G-CSF) compared with filgrastim (non-glycosylated G-CSF; P = 0.002). CD34+ cell yields were also measured halfway through the aphaeresis procedure. This was found to be highly predictive of final yield and facilitated distribution of the stem cell product to other centres. The observation that CD34+ yields did not decline in the second half compared with the first half of aphaeresis suggests that the circulating cell numbers are not static.